The goal of this proposal is to elucidate the physiological role of the novel adaptor protein HSH2 (JHematopoietic SH2 protein) in regulation of B lymphocyte biology. Studies conducted in our laboratory have demonstrated that HSH2 is expressed at low basal levels in splenic B cells and that its expression is induced in response to stimuli that bind to distinct families of receptors that promote survival and differentiation, including CD40L, IL-4, LPS, CpG DMA and BLyS (BAFF). Up-regulation of HSH2 expression was shown to be dependent on activation of NF-icB and correlates with initiation of a survival responsethat includes up-.regulation of Bcl-Xi.. Retroviral-mediated expression of HSH2 in the WEHI-231 B cell line, which undergoes apoptosis in response to BCR ligation, was observed to enhance survival and mitochondrial stability. Similarly, enhanced survival of WEHI-231 cells in response to CD40-mediated signaling directly correlated with up-regulation of HSH2 expression. Although HSH2 does not significantly alter BCR-proximal signal transduction, it was observed to maintain mitochondrial stability and this correlated with its ability to block up-regulation of Bim in response to BCR signaling. Moreover, HSH2 was found to interact with the anti-apoptotic protein HAX-1, which possessesa membrane-spanning region that targets it to the outer mitochondrial membrane. Preliminary studies have shown that the interaction between HSH2 and HAX-1 is important for the anti-apoptotic activity of HSH2. Therefore, HSH2 and HAX-1 may function together to regulate mitochondrial integrity and cell survival. To further elucidate the physiological role of HSH2 in regulation of B lymphocyte survival/differentiation, three specific aims have been proposed that will: 1) determine the physiological role of HSH2 in B lymphocyte development, homeostasis and immune function; 2) elucidate the role that HSH2 plays in regulating Bim expression in response to co-stimulation; and 3) determine the functional importance of the interaction between HSH2 and HAX-1 in regulating mitochondrial stability. Because HSH2 expression is induced in response to many of the key stimuli that are known to promote B cell survival and differentiation, this adaptor protein is likely to play an important role in regulating B cell homeostasis and immune function. Therefore, these studies will provide novel insight into the molecular mechanisms that maintain the balance between B lymphocyte survival and death leading to differentiation into humoral effector cells and will provide insight into the etiology and progression of diseases associated with aberrant B cell function, including cancer and autoimmunity.